This is a supplemental application, related to Research Grant No. AI-15145-03, which is now in its first year of a five-year project period. This project was summarized in our original application as follows: This project is concerned with cellular regulatory factors affectng DNA replication, particularly those pertaining to the enzymology and control of DNA precursor synthesis. Several lines of evidence indicate that the enzymes of deoxyribonucleotide biosynthesis interact to form a specific complex, which is juxtaposed with the DNA replication apparatus. The action of this complex can compartmentalize DNA precurosors and maintain high concentrations at the replication fork. Using the well-characterized bacteriophage T6, we hope to further explore this model. We shall attempt to purify the complex from T4-infected Escherichia coli and to determine protein-protein relationships among constituent enzymes by use of crosslinking reagents, fluorescence measurements, and attempted reconstitution. We shall seek evidence that the complex is associated with elements of the DNA replication apparatus. We shall study elements of the molecular properties of the enzymes involved and regulation of the genes which encode them. Finally, we shall study aspects of DNA precursor metabolism in T7 phage infection -aspects which should deepen our knowledge of how the enzymes of deoxyribonucleotide biosynthesis are organized intracellularly. The project remains essentially unchanges, except for two additional sub-projects: We describe methods for estimation of the effective concentrations of deoxyribonucleoside triphosphates at replication forks in intact cells. Also the complex we are studying is controlled in an unusual manner which we hope to elucidate.